This study investigated nuclear markers in Coptotermes formosanus Shiraki and Coptotermes gestroi (Wasmann) (Isoptera: Rhinotermitidae) that can be used as a diagnostic tool to detect F1 hybrids from field samples. Six microsatellite markers were compatible for both parental species and hybrid termites and were optimized so that a standard gene library can be built for the south Florida Coptotermes populations.